Project Summary In normal babies, plasma lipid levels remain relatively low until birth. Thereafter, they rise dramatically with the onset of breast or formula feeding. By the time a baby is born full term, his or her cardiomyocytes have generated the biochemical machinery to metabolize long chain fatty acids for generating ATP through ?- oxidation and oxidative phosphorylation and they are no longer susceptible for lipid toxicity. However, it appears that high levels of lipid can be toxic to immature cells. Unfortunately, there are no studies of lipotoxicity in immature cardiomyocytes. This gap is important for two reasons: 1) pregnant women who are obese, diabetic or have preeclampsia have higher than normal triglycerides and higher than normal free fatty acids. As a consequence, the cord blood of babies also has elevated levels of these lipids. 2) premature babies are often given a commercial lipid preparation with high levels of fatty acids and other complex lipids. In both cases, immature heart cells are exposed to pathologically high lipid levels. This application is designed to determine the vulnerability of developing ovine cardiomyocytes to lipid levels seen in pregnant women with compromising conditions. The purpose of this application is to understand two understudied features of heart development: 1) How cardiomyocyte metabolism can be detrimentally affected by its lipid environment before and after birth and 2) how the metabolic maturation process in cardiomyocytes is regulated by three powerful influences that remain unstudied in the context of lipid metabolism. Thus, our global hypothesis is that 1) immature cardiomyocytes are more vulnerable to abnormal elevations in fetal lipid levels than postnatal myocytes and 2) the maturation of cardiomyocytes is augmented by normal reductions in the expression of the transcription factor, Meis1, and elevations of thyroid hormone levels in the fetal blood and is suppressed by placental insufficiency. Aim 1 will determine the toxic levels of the saturated long chain fatty acid, palmitic acid (PA) and the mono-unsaturated fatty acid, oleic acid (OA) for fetal cardiomyocytes taken from sheep at 100d, 135d, and 14d neonates. In addition, the ability of these cells to take up and store fluorescent long chain fatty acid analogue, BODIPY C-12, will also be determined before and after exposure to high levels of PA and OA. Aim 2 will determine the role of Meis1 in regulating the metabolic transition from glycolysis to oxidative phosphorylation in maturing cardiomyocytes, Aim 3 will determine the degree to which thyroid hormone promotes maturation of cardiomyocytes and resistance to lipotoxicity and Aim 4 will determine the degree to which placental insufficiency stunts maturity of cardiomyocytes and promotes their vulnerability for lipotoxicity. Findings will provide the basis for new studies on human infants to help provide the most helpful fuel mixes for babies who need nutrition therapy and who are born with toxic levels of plasma long chain fatty acids.